The Development of Kisspeptin Circuits in the Mammalian Brain - Kisspeptin Signaling in Reproductive Biology - page 235

correct adult sex steroidal milieu [ 94 ]. Thus, the reduction of these fi bers in non-E 2
treated hpg (and KERKO) mice is likely due to an activational defect (i.e., absence
of circulating sex steroids). However, caution must be used in interpreting fi ber data
since the origin of the fi bers cannot always be determined.
The hpg and KERKO mouse models have recently been utilized to shed more
light on the role of gonadal sex steroids in the development of ARC Kiss1 neurons.
First, kisspeptin cell bodies are more visible in the ARC of adult hpg mice than WT
mice [ 94 ]. This difference in detectable kisspeptin cells is likely due to the lack of
obstruction by dense kisspeptin fi bers in hpg mice, as well as a possible increase in
ARC kisspeptin synthesis in hpg mice lacking steroid negative feedback (Fig. 11.8 ).
Since differences in ARC kisspeptin-ir expression and cell number were not easily
measurable by IHC due to the obstructive overlaying fi ber network, Kiss1 mRNA
levels were examined instead using ISH. Both the number of Kiss1 neurons and the
density of silver grains were increased in the ARC of adult hpg mice compared to
WT littermates, suggesting an increased level of kisspeptin synthesis in hpg mice,
perhaps due to less steroid negative feedback. Interestingly, at PND 11, only a small
difference in Kiss1 levels was present between WT and hpg littermates (as mea-
sured by RT-PCR), but by PND 31 this difference was much larger, with dramati-
cally higher ARC Kiss1 expression in hpg mice. This indicates that, at PND 11,
there may be non-gonadal hormone factors inhibiting Kiss1 in the ARC, but by
PND 31 this gonadal-independent mechanism is reduced (or absent). These hpg
data therefore suggest that changes in gonad-independent factors may induce ARC
Kiss1 expression around puberty.
The robust ARC Kiss1 expression seen in cell bodies of adult hpg and ArKO
mice may be similar to the hypertrophied Kiss1 neurons seen in postmenopausal
women [ 43 , 127 ]; in all cases, the elevated Kiss1 levels may refl ect a lack of nega-
tive feedback of sex steroids. Finally, female KERKO mice show decreased kiss-
peptin- ir fi bers in the ARC at PND 16, 26, and 35, similar to hpg mice [ 10 ].
Surprisingly, unlike hpg mice, adult KERKO mice did not display the high-
expressing kisspeptin-ir cell bodies in the ARC, even though the same kisspeptin
antibody was used in both studies. However, when mRNA was analyzed by qPCR
in the mediobasal hypothalamus (which includes the ARC), Kiss1 levels in KERKO
mice were in fact increased relative to WT levels, similar to data from hpg mice.
This unexpected difference between protein and mRNA levels in the KERKO mice
has not yet been explained.
Recent work has shown that during the prepubertal period of mice, the Kiss1
response to GDX is sexually dimorphic. While prepubertal females display elevated
Kiss1 expression in the ARC, along with increased LH levels, several days after
GDX, prepubertal male mice do not (Fig. 11.9 ). Instead, if prepubertal male mice
are GDX at PND 14, no increase is seen in ARC Kiss1 cell number or serum LH
levels when measured 2-4 days later (PND 16-18) [ 44 ]. Because this prepubertal
sex difference is only observable after the removal of gonadal hormones (there is no
sex differences in ARC Kiss1 cell number at this age in gonadally intact mice),
these results suggest a sexually dimorphic difference in the regulation of ARC Kiss1
cells at this point in development: Kiss1 /LH levels in PND 16-18 females appear to